Mechanism of CD8 regulation of natural killer cell biology

项目摘要 血液系统恶性肿瘤，如急性髓系白血病(AML)，在临床上仍然具有挑战性 对传统化疗方法无效。对许多患者来说，造血细胞移植(HCT)是 唯一的根治性治疗，其中部分疗效是由捐赠者的自然杀伤(NK)细胞靶向 受者的白血病细胞。AML患者的自然杀伤细胞过继治疗是一种很有前途的方法 以改善患者的预后。Fehniger实验室的研究确定了一种策略，可以诱导类似记忆的 (Ml)使用IL-12、IL-15和IL-18刺激分离的NK细胞的表型，显示增强 体外和体内的增殖、细胞毒性和持久性。应用ML-NK细胞进行的一期研究 复发/难治性AML患者显示约50%的患者完全缓解， 虽然导致治疗失败的机制还不清楚。 多维免疫相关分析发现供者NK细胞与 NK细胞表面CD8表达与治疗疗效的关系然而，CD8在人类常规和ML中的作用 NK细胞仍不清楚。 这项拟议的项目旨在了解CD8表达影响传统 和ML NK细胞生物学，对于了解NK细胞在癌症、感染、 和自身免疫力。初步数据显示CD8+NK细胞的增殖能力减弱 与CD8-NK细胞在体外和体内进行比较。我们推测供者NK细胞上CD8的表达 对整体NK细胞对AML的反应有负面影响。本提案的目标1侧重于确定 CD8+常规NK细胞和ML NK细胞抑制增殖的机制及其后续的 对体内肿瘤控制的影响。具体地说，端粒长度、染色质可及性和 将调查新陈代谢的差异。目标2侧重于定义CD8分子本身的功能 通过基于CRISPR-Cas9的高效击倒，以及随后对传统和ML的影响 NK细胞的细胞毒作用、信号传递和存活率。总而言之，这些结果将进一步指导未来的NK细胞治疗 设计并加强我们对CD8及其对NK细胞生物学基本方面的贡献的理解。

Project Summary Hematologic malignancies such as acute myeloid leukemia (AML) remain clinically challenging and refractory to traditional chemotherapy approaches. For many patients, hematopoietic cell transplant (HCT) is the only curative treatment, whereby part of the efficacy is driven by donor natural killer (NK) cells that target the recipient’s leukemic cells. Natural killer cell adoptive therapy for AML patients represent a promising approach for improving patient outcomes. Work done in the Fehniger lab has defined a strategy to induce a memory-like (ML) phenotype of isolated NK cells using a stimulation with IL-12, IL-15, and IL-18 that demonstrates enhanced proliferation, cytotoxicity, and persistence both in vitro and in vivo. A phase 1 study using ML NK cells in relapsed/refractory AML patients showed approximately 50% of patients achieving a complete remission, although the mechanisms driving treatment failure are not clearly understood. Multidimensional immune correlative analysis of donor NK cells identified a negative association between CD8 expression on NK cells and treatment response. However, the role of CD8 on human conventional and ML NK cells remains unknown. The proposed project aims to understand the mechanism by which CD8 expression impacts conventional and ML NK cell biology, and has broad implications for understanding NK cell responses in cancer, infection, and autoimmunity. Preliminary data demonstrate that CD8+ NK cells have a diminished proliferative capacity compared to CD8- NK cells both in vitro and in vivo. We hypothesize that CD8 expression on donor NK cells negatively impacts overall NK cell responses against AML. Aim 1 of this proposal focuses on determining the mechanisms by which CD8+ conventional and ML NK cells have compromised proliferation, and the subsequent impact on tumor control in vivo. Specifically, the contributions of telomere length, chromatin accessibility, and metabolic differences will be investigated. Aim 2 focuses on defining the function of the CD8 molecule itself through highly efficient CRISPR-Cas9 based knockdown, and the subsequent impact on conventional and ML NK cell cytotoxicity, signaling, and survival. Together, these results will further inform future NK cell therapy designs and enhance our understanding of CD8 and its contributions to fundamental aspects of NK cell biology.